The present invention concerns the hepatitis B virus genome with a vaccine-induced mutation at amino acid residue 145 (Glycine to Arginine) within the major surface antigen, its nucleotide sequence, the deduced four major protein sequences, antigen, antibody, detection systems, development of effective vaccines, and antiviral agents.
The hepatitis B virus was first discovered in 1963 as a human virus that is transmitted parenterally. Although these viruses are not particularly cytotoxic and do not lead to massive cell death, they have been the cause of a major infectious disease affecting both adults and young children worldwide. The presence of hepatitis B surface antigen has served as the main detection marker for carriers of hepatitis B virus, and thus, possibly those at risk of transmitting the virus. Conversely, the occurrence of an anti-surface antigen antibody indicates an immune response which would lead to eventual recovery. Stimulation of such immune response has been greatly helped by the currently licensed hepatitis B vaccines developed by Merck Sharpe & Dohme. These vaccines contain the major surface antigen in either the natural (plasma-derived) or the recombinant (purified from yeast cells) form, and have proven safe and effective in neutralizing the hepatitis B virus. In Singapore, the active vaccination program at a national scale has resulted in a significant decrease of acute hepatitis B infection and the incidence of primary hepatocellular carcinoma. This decrease has in turn been associated with an increased immunity in the population.
The major antigenic epitope of hepatitis B virus is a highly conserved region spanning 23 amino acid residues and located from amino acid position 124 to 147 of the major surface antigen. This small region designated as the group specific determinant “a” is found in all subtypes and isolates of hepatitis B viral genomes. Its antigenic properties seem due to its proposed double loop structure, to which the vaccine-induced neutralizing antibody binds.
In contrast to random mutations introduced into hepatitis B viral genomes during viral replication by the proof-reading defective reverse transcriptase, mutations induced following vaccination occur mainly in the “a” epitope of the major surface antigen. These mutant viruses are of particular interest since they show reduced affinity to the neutralizing antibody and therefore are able to replicate independently. Among these vaccine-escape mutants, the mutation at amino acid residue 145 (from Glycine to Arginine) in the second loop of the major surface antigen is the most significant because it is stable, results in conformational changes of the “a” epitope and has been reported worldwide in North America, Europe, Japan and Southeast Asia. In Singapore, for example, such mutants are the most frequent variant following vaccination. Twelve infectious variants among 41 breakthroughs have been identified as having an arginine mutation at amino acid residue 145 of the major surface antigen. There is evidence of vertical transmission from one of the 12 variants and this variant has also been associated with active liver disease. Significantly, some of these variants are now found in random asymptomatic adult population.
The occurrence of this replicative vaccine-induced mutant and its ability to escape detection using standard reagents is of grave concern because it has resulted in the development of acute hepatitis B in Italy and Singapore. This situation therefore requires the urgent development of specific detection systems, as well as, effective prophylactic vaccines and antiviral agents. Determination of the nucleotide sequence of this vaccine-induced mutant virus constitutes the first step towards these aims and will certainly be helpful for the various above-mentioned developments.